Treating cancer with a combination of a pd-1 antagonist and dinaciclib

ABSTRACT

The present disclosure describes combination therapies comprising an antagonist of Programmed Death 1 receptor (PD-1) and the CDK inhibitor dincaciclib, and the use of the combination therapies for the treatment of cancer, and in particular for treating cancers that express PD-L1.

FIELD OF THE INVENTION

The present invention relates to combination therapies useful for thetreatment of cancer. In particular, the invention relates to acombination therapy which comprises an antagonist of a Programmed Death1 protein (PD-1) and dinaciclib, which is a pan cyclin-dependent kinase(CDK) inhibitor.

REFERENCE TO SEQUENCE LISTING SUBMITTED ELECTRONICALLY

The sequence listing of the present application is submittedelectronically via EFS-Web as an ASCII formatted sequence listing with afile name “23572USPCTSequence.txt”, creation date of Jul. 15, 2014, anda size of 31 KB. This sequence listing submitted via EFS-Web is part ofthe specification and is herein incorporated by reference in itsentirety.

BACKGROUND OF THE INVENTION

PD-1 is recognized as an important player in immune regulation and themaintenance of peripheral tolerance. PD-1 is moderately expressed onnaive T, B and NKT cells and up-regulated by T/B cell receptor signalingon lymphocytes, monocytes and myeloid cells (1).

Two known ligands for PD-1, PD-L1 (B7-H1) and PD-L2 (B7-DC), areexpressed in human cancers arising in various tissues. In large samplesets of e.g. ovarian, renal, colorectal, pancreatic, liver cancers andmelanoma, it was shown that PD-L1 expression correlated with poorprognosis and reduced overall survival irrespective of subsequenttreatment (2-13). Similarly, PD-1 expression on tumor infiltratinglymphocytes was found to mark dysfunctional T cells in breast cancer andmelanoma (14-15) and to correlate with poor prognosis in renal cancer(16). Thus, it has been proposed that PD-L1 expressing tumor cellsinteract with PD-1 expressing T cells to attenuate T cell activation andevasion of immune surveillance, thereby contributing to an impairedimmune response against the tumor.

Several monoclonal antibodies that inhibit the interaction between PD-1and one or both of its ligands PD-L1 and PD-L2 are in clinicaldevelopment for treating cancer. It has been proposed that the efficacyof such antibodies might be enhanced if administered in combination withother approved or experimental cancer therapies, e.g., radiation,surgery, chemotherapeutic agents, targeted therapies, agents thatinhibit other signaling pathways that are disregulated in tumors, andother immune enhancing agents.

Disregulation of cell cycle control is a hallmark of all human cancersand is frequently associated with aberrant activation/regulation ofcyclin-dependent kinases (CDKs). However, the CDK cascade is importantfor maintaining proper function of T cells in a context-dependentmanner. Thus, the development of CDK inhibitors (CKDIs) as anti-canceragents has been complicated by immune cell toxicities and the consequentpotential for immunosuppressive effects (17). Dinaciclib, a pan-CDKinhibitor which selectively inhibits CDK1, CDK2, CDK5, and CDK9, hasbeen investigated as a potential therapy in a variety of cancers, withneutropenia being the most common dose-limiting toxicity observed inclinical trials (17, 18).

SUMMARY OF THE INVENTION

The present invention is based, in part, on the surprising finding thatconcurrent administration of dinaciclib and a murinized anti-mouse PD-1antibody to tumor-bearing mice resulted in significantly higheranti-tumor efficacy compared to either agent alone. This finding wasunexpected because the known activity of dinaciclib in potentlyinhibiting transcription and cell proliferation was predicted tocounteract the effectiveness of anti-PD-1 therapy, which is believed tolargely involve the activation and proliferation of T cells present inand recruited to a tumor.

Thus, in one embodiment, the invention provides a method for treating acancer in an individual comprising administering to the individual acombination therapy which comprises a PD-1 antagonist and a dinaciclibcompound.

In another embodiment, the invention provides a medicament comprising aPD-1 antagonist for use in combination with a dinaciclib compound fortreating a cancer.

In yet another embodiment, the invention provides a medicamentcomprising a dinaciclib compound for use in combination with a PD-1antagonist for treating a cancer.

Other embodiments provide use of a PD-1 antagonist in the manufacture ofmedicament for treating a cancer in an individual when administered incombination with a dinaciclib compound and use of a dinaciclib compoundin the manufacture of a medicament for treating a cancer in anindividual when administered in combination with a PD-1 antagonist.

In a still further embodiment, the invention provides use of a PD-1antagonist and a dinaciclib compound in the manufacture of medicamentsfor treating a cancer in an individual. In some preferred embodiments,the medicaments comprise a kit, and the kit also comprises a packageinsert comprising instructions for using the PD-1 antagonist incombination with a dinaciclib compound to treat a cancer in anindividual.

In all of the above treatment method, medicaments and uses, the PD-1antagonist inhibits the binding of PD-L1 to PD-1, and preferably alsoinhibits the binding of PD-L2 to PD-1. In some preferred embodiments ofthe above treatment method, medicaments and uses, the PD-1 antagonist isa monoclonal antibody, or an antigen binding fragment thereof, whichspecifically binds to PD-1 or to PD-L1 and blocks the binding of PD-L1to PD-1. In one particularly preferred embodiment, the PD-1 antagonistis an anti-PD-1 antibody which comprises a heavy chain and a lightchain, and wherein the heavy and light chains comprise the amino acidsequences shown in FIG. 6 (SEQ ID NO:21 and SEQ ID NO:22).

In all of the above embodiments of the treatment method, medicaments anduses, the dinaciclib compound is the compound of Formula I

or a pharmaceutically acceptable salt of the compound of Formula I.

In some embodiments of the above treatment method, medicaments and usesof the invention, the individual is a human and the cancer is a solidtumor and in some preferred embodiments, the solid tumor is bladdercancer, breast cancer, clear cell kidney cancer, head/neck squamous cellcarcinoma, lung squamous cell carcinoma, malignant melanoma,non-small-cell lung cancer (NSCLC), ovarian cancer, pancreatic cancer,prostate cancer, renal cell cancer, small-cell lung cancer (SCLC) ortriple negative breast cancer. In some preferred embodiments, the canceris ipilimumab-naïve advanced melanoma and while in other preferredembodiments, the human has ipilimumab-refractory advanced melanoma.

In other embodiments of the above treatment method, medicaments and usesof the invention, the individual is a human and the cancer is a Hememalignancy and in some preferred embodiments, the Heme malignancy isacute lymphoblastic leukemia (ALL), acute myeloid leukemia (AML),chronic lymphocytic leukemia (CLL), chronic myeloid leukemia (CML),diffuse large B-cell lymphoma (DLBCL), EBV-positive DLBCL, primarymediastinal large B-cell lymphoma, T-cell/histiocyte-rich large B-celllymphoma, follicular lymphoma, Hodgkin's lymphoma (HL), mantle celllymphoma (MCL), multiple myeloma (MM), myeloid cell leukemia-1 protein(Mc1-1), myelodysplastic syndrome (MDS), non-Hodgkin's lymphoma (NHL),or small lymphocytic lymphoma (SLL).

Also, in preferred embodiments of any of the above treatment method,medicaments and uses, the cancer expresses one or both of PD-L1 andPD-L2. In particularly preferred embodiments, PD-L1 expression iselevated in the cancer.

In one particularly preferred embodiment of the above treatment method,medicaments and uses, the individual is a human and the cancer ischronic lymphocytic leukemia (CLL) that expresses human PD-L1.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 shows amino acid sequences of the light chain and heavy chainCDRs for an exemplary anti-PD-1 monoclonal antibody useful in thepresent invention (SEQ ID NOs:1-6).

FIG. 2 shows amino acid sequences of the light chain and heavy chainCDRs for another exemplary anti-PD-1 monoclonal antibody useful in thepresent invention (SEQ ID NOs:7-12).

FIG. 3 shows amino acid sequences of the heavy chain variable region andfull length heavy chain for an exemplary anti-PD-1 monoclonal antibodyuseful in the present invention (SEQ ID NO:13 and SEQ ID NO:14).

FIG. 4 shows amino acid sequences of alternative light chain variableregions for an exemplary anti-PD-1 monoclonal antibody useful in thepresent invention (SEQ ID NOs:15-17).

FIG. 5 shows amino acid sequences of alternative light chains for anexemplary anti-PD-1 monoclonal antibody useful in the present invention(SEQ ID NOs:18-20).

FIG. 6 shows amino acid sequences of the heavy and light chains forMK-3475 (SEQ ID NOs. 21 and 22, respectively).

FIG. 7 shows amino acid sequences of the heavy and light chains fornivolumab (SEQ ID NOs. 23 and 24, respectively).

FIG. 8 illustrates the anti-tumor effect of concurrent administration ofa PD-1 antagonist and dinaciclib is superior to monotherapy with eitheragent alone in tumor-bearing mice, with FIG. 8A showing the mean tumorvolume at various days during treatment with a control, a murineanti-mouse PD-1 mAb (Anti-PD1), dinaciclib, or both Anti-PD1 anddinaciclib, and FIG. 8B showing the tumor volume values for individualmice in each treatment group on the first day of treatment (left graph,Day 0) or after 25 days of treatment (right graph, Day 25). Experimentaldetails are described in Example 1 below.

DETAILED DESCRIPTION Abbreviations

Throughout the detailed description and examples of the invention thefollowing abbreviations will be used:

CDR Complementarity determining region

CHO Chinese hamster ovary

FFPE formalin-fixed, paraffin-embedded

FR Framework region

IgG Immunoglobulin G

IHC Immunohistochemistry or immunohistochemical

Q2W One dose every two weeks

Q3W One dose every three weeks

VH Immunoglobulin heavy chain variable region

VK Immunoglobulin kappa light chain variable region

I. DEFINITIONS

So that the invention may be more readily understood, certain technicaland scientific terms are specifically defined below. Unless specificallydefined elsewhere in this document, all other technical and scientificterms used herein have the meaning commonly understood by one ofordinary skill in the art to which this invention belongs.

As used herein, including the appended claims, the singular forms ofwords such as “a,” “an,” and “the,” include their corresponding pluralreferences unless the context clearly dictates otherwise.

“Administration” and “treatment,” as it applies to an animal, human,experimental subject, cell, tissue, organ, or biological fluid, refersto contact of an exogenous pharmaceutical, therapeutic, diagnosticagent, or composition to the animal, human, subject, cell, tissue,organ, or biological fluid. Treatment of a cell encompasses contact of areagent to the cell, as well as contact of a reagent to a fluid, wherethe fluid is in contact with the cell. “Administration” and “treatment”also means in vitro and ex vivo treatments, e.g., of a cell, by areagent, diagnostic, binding compound, or by another cell. The term“subject” includes any organism, preferably an animal, more preferably amammal (e.g., rat, mouse, dog, cat, rabbit) and most preferably a human.

As used herein, the term “antibody” refers to any form of antibody thatexhibits the desired biological or binding activity. Thus, it is used inthe broadest sense and specifically covers, but is not limited to,monoclonal antibodies (including full length monoclonal antibodies),polyclonal antibodies, multispecific antibodies (e.g., bispecificantibodies), humanized, fully human antibodies, chimeric antibodies andcamelized single domain antibodies. “Parental antibodies” are antibodiesobtained by exposure of an immune system to an antigen prior tomodification of the antibodies for an intended use, such as humanizationof an antibody for use as a human therapeutic.

In general, the basic antibody structural unit comprises a tetramer.Each tetramer includes two identical pairs of polypeptide chains, eachpair having one “light” (about 25 kDa) and one “heavy” chain (about50-70 kDa). The amino-terminal portion of each chain includes a variableregion of about 100 to 110 or more amino acids primarily responsible forantigen recognition. The carboxy-terminal portion of the heavy chain maydefine a constant region primarily responsible for effector function.Typically, human light chains are classified as kappa and lambda lightchains. Furthermore, human heavy chains are typically classified as mu,delta, gamma, alpha, or epsilon, and define the antibody's isotype asIgM, IgD, IgG, IgA, and IgE, respectively. Within light and heavychains, the variable and constant regions are joined by a “J” region ofabout 12 or more amino acids, with the heavy chain also including a “D”region of about 10 more amino acids. See generally, FundamentalImmunology Ch. 7 (Paul, W., ed., 2nd ed. Raven Press, N.Y. (1989).

The variable regions of each light/heavy chain pair form the antibodybinding site. Thus, in general, an intact antibody has two bindingsites. Except in bifunctional or bispecific antibodies, the two bindingsites are, in general, the same.

Typically, the variable domains of both the heavy and light chainscomprise three hypervariable regions, also called complementaritydetermining regions (CDRs), which are located within relativelyconserved framework regions (FR). The CDRs are usually aligned by theframework regions, enabling binding to a specific epitope. In general,from N-terminal to C-terminal, both light and heavy chains variabledomains comprise FR1, CDR1, FR2, CDR2, FR3, CDR3 and FR4. The assignmentof amino acids to each domain is, generally, in accordance with thedefinitions of Sequences of Proteins of Immunological Interest, Kabat,et al.; National Institutes of Health, Bethesda, Md.; 5^(th) ed.; NIHPubl. No. 91-3242 (1991); Kabat (1978) Adv. Prot. Chem. 32:1-75; Kabat,et al., (1977) J. Biol. Chem. 252:6609-6616; Chothia, et al., (1987) JMol. Biol. 196:901-917 or Chothia, et al., (1989) Nature 342:878-883.

As used herein, the term “hypervariable region” refers to the amino acidresidues of an antibody that are responsible for antigen-binding. Thehypervariable region comprises amino acid residues from a“complementarity determining region” or “CDR” (i.e. CDRL1, CDRL2 andCDRL3 in the light chain variable domain and CDRH1, CDRH2 and CDRH3 inthe heavy chain variable domain). See Kabat et al. (1991) Sequences ofProteins of Immunological Interest, 5th Ed. Public Health Service,National Institutes of Health, Bethesda, Md. (defining the CDR regionsof an antibody by sequence); see also Chothia and Lesk (1987) J. Mol.Biol. 196: 901-917 (defining the CDR regions of an antibody bystructure). As used herein, the term “framework” or “FR” residues refersto those variable domain residues other than the hypervariable regionresidues defined herein as CDR residues.

As used herein, unless otherwise indicated, “antibody fragment” or“antigen binding fragment” refers to antigen binding fragments ofantibodies, i.e. antibody fragments that retain the ability to bindspecifically to the antigen bound by the full-length antibody, e.g.fragments that retain one or more CDR regions. Examples of antibodybinding fragments include, but are not limited to, Fab, Fab′, F(ab′)₂,and Fv fragments; diabodies; linear antibodies; single-chain antibodymolecules, e.g., sc-Fv; nanobodies and multispecific antibodies formedfrom antibody fragments.

An antibody that “specifically binds to” a specified target protein isan antibody that exhibits preferential binding to that target ascompared to other proteins, but this specificity does not requireabsolute binding specificity. An antibody is considered “specific” forits intended target if its binding is determinative of the presence ofthe target protein in a sample, e.g. without producing undesired resultssuch as false positives. Antibodies, or binding fragments thereof,useful in the present invention will bind to the target protein with anaffinity that is at least two fold greater, preferably at least tentimes greater, more preferably at least 20-times greater, and mostpreferably at least 100-times greater than the affinity with non-targetproteins. As used herein, an antibody is said to bind specifically to apolypeptide comprising a given amino acid sequence, e.g. the amino acidsequence of a mature human PD-1 or human PD-L1 molecule, if it binds topolypeptides comprising that sequence but does not bind to proteinslacking that sequence.

“Chimeric antibody” refers to an antibody in which a portion of theheavy and/or light chain is identical with or homologous tocorresponding sequences in an antibody derived from a particular species(e.g., human) or belonging to a particular antibody class or subclass,while the remainder of the chain(s) is identical with or homologous tocorresponding sequences in an antibody derived from another species(e.g., mouse) or belonging to another antibody class or subclass, aswell as fragments of such antibodies, so long as they exhibit thedesired biological activity.

“Human antibody” refers to an antibody that comprises humanimmunoglobulin protein sequences only. A human antibody may containmurine carbohydrate chains if produced in a mouse, in a mouse cell, orin a hybridoma derived from a mouse cell. Similarly, “mouse antibody” or“rat antibody” refer to an antibody that comprises only mouse or ratimmunoglobulin sequences, respectively.

“Humanized antibody” refers to forms of antibodies that containsequences from non-human (e.g., murine) antibodies as well as humanantibodies. Such antibodies contain minimal sequence derived fromnon-human immunoglobulin. In general, the humanized antibody willcomprise substantially all of at least one, and typically two, variabledomains, in which all or substantially all of the hypervariable loopscorrespond to those of a non-human immunoglobulin and all orsubstantially all of the FR regions are those of a human immunoglobulinsequence. The humanized antibody optionally also will comprise at leasta portion of an immunoglobulin constant region (Fc), typically that of ahuman immunoglobulin. The prefix “hum”, “hu” or “h” is added to antibodyclone designations when necessary to distinguish humanized antibodiesfrom parental rodent antibodies. The humanized forms of rodentantibodies will generally comprise the same CDR sequences of theparental rodent antibodies, although certain amino acid substitutionsmay be included to increase affinity, increase stability of thehumanized antibody, or for other reasons.

The terms “cancer”, “cancerous”, or “malignant” refer to or describe thephysiological condition in mammals that is typically characterized byunregulated cell growth. Examples of cancer include but are not limitedto, carcinoma, lymphoma, leukemia, blastoma, and sarcoma. Moreparticular examples of such cancers include squamous cell carcinoma,myeloma, small-cell lung cancer, non-small cell lung cancer, glioma,hodgkin's lymphoma, non-hodgkin's lymphoma, acute myeloid leukemia(AML), multiple myeloma, gastrointestinal (tract) cancer, renal cancer,ovarian cancer, liver cancer, lymphoblastic leukemia, lymphocyticleukemia, colorectal cancer, endometrial cancer, kidney cancer, prostatecancer, thyroid cancer, melanoma, chondrosarcoma, neuroblastoma,pancreatic cancer, glioblastoma multiforme, cervical cancer, braincancer, stomach cancer, bladder cancer, hepatoma, breast cancer, coloncarcinoma, and head and neck cancer. Particularly preferred cancers thatmay be treated in accordance with the present invention include thosecharacterized by elevated expression of one or both of PD-L1 and PD-L2in tested tissue samples.

“Biotherapeutic agent” means a biological molecule, such as an antibodyor fusion protein, that blocks ligand/receptor signaling in anybiological pathway that supports tumor maintenance and/or growth orsuppresses the anti-tumor immune response.

“CDR” or “CDRs” as used herein means complementarity determiningregion(s) in a immunoglobulin variable region, defined using the Kabatnumbering system, unless otherwise indicated

“Chemotherapeutic agent” is a chemical compound useful in the treatmentof cancer. Classes of chemotherapeutic agents include, but are notlimited to: alkylating agents, antimetabolites, kinase inhibitors,spindle poison plant alkaloids, cytoxic/antitumor antibiotics,topisomerase inhibitors, photosensitizers, anti-estrogens and selectiveestrogen receptor modulators (SERMs), anti-progesterones, estrogenreceptor down-regulators (ERDs), estrogen receptor antagonists,leutinizing hormone-releasing hormone agonists, anti-androgens,aromatase inhibitors, EGFR inhibitors, VEGF inhibitors, anti-senseoligonucleotides that that inhibit expression of genes implicated inabnormal cell proliferation or tumor growth. Chemotherapeutic agentsuseful in the treatment methods of the present invention includecytostatic and/or cytotoxic agents.

“Clothia” as used herein means an antibody numbering system described inAl-Lazikani et al., JMB 273:927-948 (1997).

“Conservatively modified variants” or “conservative substitution” refersto substitutions of amino acids in a protein with other amino acidshaving similar characteristics (e.g. charge, side-chain size,hydrophobicity/hydrophilicity, backbone conformation and rigidity,etc.), such that the changes can frequently be made without altering thebiological activity or other desired property of the protein, such asantigen affinity and/or specificity. Those of skill in this artrecognize that, in general, single amino acid substitutions innon-essential regions of a polypeptide do not substantially alterbiological activity (see, e.g., Watson et al. (1987) Molecular Biologyof the Gene, The Benjamin/Cummings Pub. Co., p. 224 (4th Ed.)). Inaddition, substitutions of structurally or functionally similar aminoacids are less likely to disrupt biological activity. Exemplaryconservative substitutions are set forth in Table 3.

TABLE 1 Exemplary Conservative Amino Acid Substitutions Original residueConservative substitution Ala (A) Gly; Ser Arg (R) Lys; His Asn (N) Gln;His Asp (D) Glu; Asn Cys (C) Ser; Ala Gln (Q) Asn Glu (E) Asp; Gln Gly(G) Ala His (H) Asn; Gln Ile (I) Leu; Val Leu (L) Ile; Val Lys (K) Arg;His Met (M) Leu; Ile; Tyr Phe (F) Tyr; Met; Leu Pro (P) Ala Ser (S) ThrThr (T) Ser Trp (W) Tyr; Phe Tyr (Y) Trp; Phe Val (V) Ile; Leu

“Consists essentially of,” and variations such as “consist essentiallyof” or “consisting essentially of,” as used throughout the specificationand claims, indicate the inclusion of any recited elements or group ofelements, and the optional inclusion of other elements, of similar ordifferent nature than the recited elements, that do not materiallychange the basic or novel properties of the specified dosage regimen,method, or composition. As a non-limiting example, a PD-1 antagonistthat consists essentially of a recited amino acid sequence may alsoinclude one or more amino acids, including substitutions of one or moreamino acid residues, which do not materially affect the properties ofthe binding compound.

“Diagnostic anti-PD-L monoclonal antibody” means a mAb whichspecifically binds to the mature form of the designated PD-L (PD-L1 orPDL2) that is expressed on the surface of certain mammalian cells. Amature PD-L lacks the presecretory leader sequence, also referred to asleader peptide The terms “PD-L” and “mature PD-L” are usedinterchangeably herein, and shall be understood to mean the samemolecule unless otherwise indicated or readily apparent from thecontext.

As used herein, a diagnostic anti-human PD-L1 mAb or an anti-hPD-L1 mAbrefers to a monoclonal antibody that specifically binds to mature humanPD-L1. A mature human PD-L1 molecule consists of amino acids 19-290 ofthe following sequence:

(SEQ ID NO: 25) MRIFAVFIFMTYWHLLNAFTVTVPKDLYVVEYGSNMTIECKFPVEKQLDLAALIVYWEMEDKNIIQFVHGEEDLKVQHSSYRQRARLLKDQLSLGNAALQITDVKLQDAGVYRCMISYGGADYKRITVKVNAPYNKINQRILVVDPVTSEHELTCQAEGYPKAEVIWTSSDHQVLSGKTTTTNSKREEKLFNVTSTLRINTTTNEIFYCTFRRLDPEENHTAELVIPELPLAHPPNERTHLVILGAILLCLGVALTFIFRLRKGRMMDVK KCGIQDTNSKKQSDTHLEET.

Specific examples of diagnostic anti-human PD-L1 mAbs useful asdiagnostic mAbs for immunohistochemistry (IHC) detection of PD-L1expression in formalin-fixed, paraffin-embedded (FFPE) tumor tissuesections are antibody 20C3 and antibody 22C3, which are described in thecopending U.S. provisional patent application 61/745,386, filed December2012. These antibodies comprise the light chain and heavy chain variableregion amino acid sequences shown in Table 2 below:

TABLE 2 Monoclonal Antibodies 20C3 and 22C320C3 Light Chain Mature Variable RegionDIVMSQSPSSLAVSAGEKVTMSCKSSQSLLNSR SEQ ID NO: 26TRKNYLAWYQQKPGQSPKLLIYWASTRESGVPD RFTGSGSGTDFTLTISSVQAEDLAVYYCQQSYDVVTFGAGTKLELK 20C3 Heavy Chain Mature Variable RegionQVQVQQSGAELAEPGASVKMSCKASGYIFTSYW SEQ ID NO: 27MHWLKQRPGQGLEWIGYINPSSDYNEYSEKFMD KATLTADKASTTAYMQLISLTSEDSAVYYCARSGWLVHGDYYFDYWGQGTTLTVSS 22C3 Light Chain Mature Variable RegionDIVMSQSPSSLAVSAGEKVTMTCKSSQSLLHTS SEQ ID NO: 28TRKNYLAWYQQKPGQSPKLLIYWASTRESGVPD RFTGSGSGTDFTLTISSVQAEDLAVYYCKQSYDVVTFGAGTKLELK 22C3 Heavy Chain Mature Variable RegionQVHLQQSGAELAKPGASVKMSCKASGYTFTSYW SEQ ID NO: 29IHWIKQRPGQGLEWIGYINPSSGYHEYNQKFID KATLTADRSSSTAYMHLTSLTSEDSAVYYCARSGWLIHGDYYFDFWGQGTTLTVSS

Another anti-human PD-L1 mAb that has been reported to be useful for IHCdetection of PD-L1 expression in FFPE tissue sections (Chen, B. J. etal., Clin Cancer Res 19: 3462-3473 (2013)) is a rabbit anti-human PD-L1mAb publicly available from Sino Biological, Inc. (Beijing, P. R. China;Catalog number 10084-R015).

“Dincaciclib compound” means the compound of Formula I, andpharmaceutically acceptable salts of the compound of Formula I. Thechemical name of dinaciclib is1-[3-ethyl-7-[[(1-oxido-3-pyridinyl)methyl]amino]pyrazolo[1,5-a]pyrimidin-5-yl]-2(S)-piperidineethanol.This compound may be synthesized as described in U.S. Pat. No.7,119,200, or any other synthetic route that will be readily apparent tothe skilled artisan.

Reference to a compound of Formula I herein is understood to includereference to salts thereof; unless otherwise indicated. The term“salt(s)”, as employed herein, denotes acidic salts formed withinorganic and/or organic acids, as well as basic salts formed withinorganic and/or organic bases. In addition, when a compound of FormulaI contains both a basic moiety, such as, but not limited to a pyridineor imidazole, and an acidic moiety, such as, but not limited to acarboxylic acid, zwitterions (“inner salts”) may be formed and areincluded within the term “salt(s)” as used herein. Pharmaceuticallyacceptable salts of the compound of Formula I may be formed, forexample, by reacting the compound of Formula I with an amount of acid orbase, such as an equivalent amount, in a medium such as one in which thesalt precipitates or in an aqueous medium followed by lyophilization.

Exemplary acid addition salts of the compound of Formula I includeacetates, ascorbates, benzoates, benzenesulfonates, bisulfates, borates,butyrates, citrates, camphorates, camphorsulfonates, fumarates,hydrochlorides, hydrobromides, hydroiodides, lactates, maleates,methanesulfonates, naphthalenesulfonates, nitrates, oxalates,phosphates, propionates, salicylates, succinates, sulfates, tartarates,thiocyanates, toluenesulfonates (also known as tosylates,) and the like.Additionally, acids which are generally considered suitable for theformation of pharmaceutically useful salts from basic pharmaceuticalcompounds are discussed, for example, by S. Berge et al, Journal ofPharmaceutical Sciences (1977) 66(1) 1-19; P. Gould, International J. ofPharmaceutics (1986) 33 201-217; Anderson et al, The Practice ofMedicinal Chemistry (1996), Academic Press, New York; and in The OrangeBook (Food & Drug Administration, Washington, D.C. on their website).These disclosures are incorporated herein by reference thereto.

Exemplary basic salts of the compound of Formula I include ammoniumsalts, alkali metal salts such as sodium, lithium, and potassium salts,alkaline earth metal salts such as calcium and magnesium salts, saltswith organic bases (for example, organic amines) such asdicyclohexylamines, t-butyl amines, and salts with amino acids such asarginine, lysine and the like. Basic nitrogen-containing groups may bequarternized with agents such as lower alkyl halides (e.g. methyl,ethyl, and butyl chlorides, bromides and iodides), dialkyl sulfates(e.g. dimethyl, diethyl, and dibutyl sulfates), long chain halides (e.g.decyl, lauryl, and stearyl chlorides, bromides and iodides), aralkylhalides (e.g. benzyl and phenethyl bromides), and others.

All such acid salts and base salts are intended to be pharmaceuticallyacceptable salts within the scope of a dinaciclib compound used in thepresent invention and all acid and base salts are considered equivalentto the free forms of the corresponding compound for purposes of theinvention.

Prodrugs of the compound of Formula I are also contemplated for use inthe methods, medicaments and uses of the present invention. The term“prodrug”, as employed herein, denotes a compound that is a drugprecursor which, upon administration to a subject, undergoes chemicalconversion by metabolic or chemical processes to yield a compound ofFormula I or a salt thereof. A discussion of prodrugs is provided in T.Higuchi and V. Stella, Pro-drugs as Novel Delivery Systems (1987) 14 ofthe A.C.S. Symposium Series, and in Bioreversible Carriers in DrugDesign, (1987) Edward B. Roche, ed., American Pharmaceutical Associationand Pergamon Press, both of which are incorporated herein by referencethereto.

“Framework region” or “FR” as used herein means the immunoglobulinvariable regions excluding the CDR regions.

“Homology” refers to sequence similarity between two polypeptidesequences when they are optimally aligned. When a position in both ofthe two compared sequences is occupied by the same amino acid monomersubunit, e.g., if a position in a light chain CDR of two different Absis occupied by alanine, then the two Abs are homologous at thatposition. The percent of homology is the number of homologous positionsshared by the two sequences divided by the total number of positionscompared ×100. For example, if 8 of 10 of the positions in two sequencesare matched or homologous when the sequences are optimally aligned thenthe two sequences are 80% homologous. Generally, the comparison is madewhen two sequences are aligned to give maximum percent homology. Forexample, the comparison can be performed by a BLAST algorithm whereinthe parameters of the algorithm are selected to give the largest matchbetween the respective sequences over the entire length of therespective reference sequences.

The following references relate to BLAST algorithms often used forsequence analysis: BLAST ALGORITHMS: Altschul, S. F., et al., (1990) J.Mol. Biol. 215:403-410; Gish, W., et al., (1993) Nature Genet.3:266-272; Madden, T. L., et al., (1996) Meth. Enzymol. 266:131-141;Altschul, S. F., et al., (1997) Nucleic Acids Res. 25:3389-3402; Zhang,J., et al., (1997) Genome Res. 7:649-656; Wootton, J. C., et al., (1993)Comput. Chem. 17:149-163; Hancock, J. M. et al., (1994) Comput. Appl.Biosci. 10:67-70; ALIGNMENT SCORING SYSTEMS: Dayhoff, M. O., et al., “Amodel of evolutionary change in proteins.” in Atlas of Protein Sequenceand Structure, (1978) vol. 5, suppl. 3. M. O. Dayhoff (ed.), pp.345-352, Natl. Biomed. Res. Found., Washington, D.C.; Schwartz, R. M.,et al., “Matrices for detecting distant relationships.” in Atlas ofProtein Sequence and Structure, (1978) vol. 5, suppl. 3.” M. O. Dayhoff(ed.), pp. 353-358, Natl. Biomed. Res. Found., Washington, D.C.;Altschul, S. F., (1991) J. Mol. Biol. 219:555-565; States, D. J., etal., (1991) Methods 3:66-70; Henikoff, S., et al., (1992) Proc. Natl.Acad. Sci. USA 89:10915-10919; Altschul, S. F., et al., (1993) J. Mol.Evol. 36:290-300; ALIGNMENT STATISTICS: Karlin, S., et al., (1990) Proc.Natl. Acad. Sci. USA 87:2264-2268; Karlin, S., et al., (1993) Proc.Natl. Acad. Sci. USA 90:5873-5877; Dembo, A., et al., (1994) Ann. Prob.22:2022-2039; and Altschul, S. F. “Evaluating the statisticalsignificance of multiple distinct local alignments.” in Theoretical andComputational Methods in Genome Research (S. Suhai, ed.), (1997) pp.1-14, Plenum, N.Y.

“Isolated antibody” and “isolated antibody fragment” refers to thepurification status and in such context means the named molecule issubstantially free of other biological molecules such as nucleic acids,proteins, lipids, carbohydrates, or other material such as cellulardebris and growth media. Generally, the term “isolated” is not intendedto refer to a complete absence of such material or to an absence ofwater, buffers, or salts, unless they are present in amounts thatsubstantially interfere with experimental or therapeutic use of thebinding compound as described herein.

“Kabat” as used herein means an immunoglobulin alignment and numberingsystem pioneered by Elvin A. Kabat ((1991) Sequences of Proteins ofImmunological Interest, 5th Ed. Public Health Service, NationalInstitutes of Health, Bethesda, Md.).

“Monoclonal antibody” or “mAb” or “Mab”, as used herein, refers to apopulation of substantially homogeneous antibodies, i.e., the antibodymolecules comprising the population are identical in amino acid sequenceexcept for possible naturally occurring mutations that may be present inminor amounts. In contrast, conventional (polyclonal) antibodypreparations typically include a multitude of different antibodieshaving different amino acid sequences in their variable domains,particularly their CDRs, which are often specific for differentepitopes. The modifier “monoclonal” indicates the character of theantibody as being obtained from a substantially homogeneous populationof antibodies, and is not to be construed as requiring production of theantibody by any particular method. For example, the monoclonalantibodies to be used in accordance with the present invention may bemade by the hybridoma method first described by Kohler et al. (1975)Nature 256: 495, or may be made by recombinant DNA methods (see, e.g.,U.S. Pat. No. 4,816,567). The “monoclonal antibodies” may also beisolated from phage antibody libraries using the techniques described inClackson et al. (1991) Nature 352: 624-628 and Marks et al. (1991) J.Mol. Biol. 222: 581-597, for example. See also Presta (2005) J. AllergyClin. Immunol. 116:731.

“Patient” or “subject” refers to any single subject for which therapy isdesired or that is participating in a clinical trial, epidemiologicalstudy or used as a control, including humans and mammalian veterinarypatients such as cattle, horses, dogs, and cats.

“PD-1 antagonist” means any chemical compound or biological moleculethat blocks binding of PD-L1 expressed on a cancer cell to PD-1expressed on an immune cell (T cell, B cell or NKT cell) and preferablyalso blocks binding of PD-L2 expressed on a cancer cell to theimmune-cell expressed PD-1. Alternative names or synonyms for PD-1 andits ligands include: PDCD1, PD1, CD279 and SLEB2 for PD-1; PDCD1L1,PDLL, B7H1, B7-4, CD274 and B7-H for PD-L1; and PDCD1L2, PDL2, B7-DC,Btdc and CD273 for PD-L2. In any of the treatment method, medicamentsand uses of the present invention in which a human individual is beingtreated, the PD-1 antagonist blocks binding of human PD-L1 to humanPD-1, and preferably blocks binding of both human PD-L1 and PD-L2 tohuman PD-1. Human PD-1 amino acid sequences can be found in NCBI LocusNo.: NP_005009. Human PD-L1 and PD-L2 amino acid sequences can be foundin NCBI Locus No.: NP_054862 and NP_079515, respectively.

PD-1 antagonists useful in the any of the treatment method, medicamentsand uses of the present invention include a monoclonal antibody (mAb),or antigen binding fragment thereof, which specifically binds to PD-1 orPD-L1, and preferably specifically binds to human PD-1 or human PD-L1.The mAb may be a human antibody, a humanized antibody or a chimericantibody, and may include a human constant region. In some embodimentsthe human constant region is selected from the group consisting of IgG1,IgG2, IgG3 and IgG4 constant regions, and in preferred embodiments, thehuman constant region is an IgG1 or IgG4 constant region. In someembodiments, the antigen binding fragment is selected from the groupconsisting of Fab, Fab′-SH, F(ab′)₂, scFv and Fv fragments.

Examples of mAbs that bind to human PD-1, and useful in the treatmentmethod, medicaments and uses of the present invention, are described inU.S. Pat. No. 7,521,051, U.S. Pat. No. 8,008,449, and U.S. Pat. No.8,354,509. Specific anti-human PD-1 mAbs useful as the PD-1 antagonistin the treatment method, medicaments and uses of the present inventioninclude: MK-3475, a humanized IgG4 mAb with the structure described inWHO Drug Information, Vol. 27, No. 2, pages 161-162 (2013) and whichcomprises the heavy and light chain amino acid sequences shown in FIG.6, nivolumab (BMS-936558), a human IgG4 mAb with the structure describedin WHO Drug Information, Vol. 27, No. 1, pages 68-69 (2013) and whichcomprises the heavy and light chain amino acid sequences shown in FIG.7; pidilizumab (CT-011, also known as hBAT or hBAT-1); and the humanizedantibodies h409A11, h409A16 and h409A17, which are described inWO2008/156712.

Examples of mAbs that bind to human PD-L1, and useful in the treatmentmethod, medicaments and uses of the present invention, are described inWO2013/019906, WO2010/077634 A1 and U.S. Pat. No. 8,383,796. Specificanti-human PD-L1 mAbs useful as the PD-1 antagonist in the treatmentmethod, medicaments and uses of the present invention include MPDL3280A,BMS-936559, MEDI4736, MSB0010718C and an antibody which comprises theheavy chain and light chain variable regions of SEQ ID NO:24 and SEQ IDNO:21, respectively, of WO2013/019906.

Other PD-1 antagonists useful in the any of the treatment method,medicaments and uses of the present invention include an immunoadhesinthat specifically binds to PD-1 or PD-L1, and preferably specificallybinds to human PD-1 or human PD-L1, e.g., a fusion protein containingthe extracellular or PD-1 binding portion of PD-L1 or PD-L2 fused to aconstant region such as an Fc region of an immunoglobulin molecule.Examples of immunoadhesion molecules that specifically bind to PD-1 aredescribed in WO2010/027827 and WO2011/066342. Specific fusion proteinsuseful as the PD-1 antagonist in the treatment method, medicaments anduses of the present invention include AMP-224 (also known as B7-DCIg),which is a PD-L2-FC fusion protein and binds to human PD-1.

In some preferred embodiments of the treatment method, medicaments anduses of the present invention, the PD-1 antagonist is a monoclonalantibody, or antigen binding fragment thereof, which comprises: (a)light chain CDRs SEQ ID NOs: 1, 2 and 3 and heavy chain CDRs SEQ ID NOs:4, 5 and 6; or (b) light chain CDRs SEQ ID NOs: 7, 8 and 9 and heavychain CDRs SEQ ID NOs: 10, 11 and 12.

In other preferred embodiments of the treatment method, medicaments anduses of the present invention, the PD-1 antagonist is a monoclonalantibody, or antigen binding fragment thereof, which specifically bindsto human PD-1 and comprises (a) a heavy chain variable region comprisingSEQ ID NO:13 or a variant thereof; and (b) a light chain variable regioncomprising an amino acid sequence selected from the group consisting ofSEQ ID NO:15 or a variant thereof; SEQ ID NO:16 or a variant thereof;and SEQ ID NO: 17 or a variant thereof. A variant of a heavy chainvariable region sequence is identical to the reference sequence excepthaving up to 17 conservative amino acid substitutions in the frameworkregion (i.e., outside of the CDRs), and preferably has less than ten,nine, eight, seven, six or five conservative amino acid substitutions inthe framework region. A variant of a light chain variable regionsequence is identical to the reference sequence except having up to fiveconservative amino acid substitutions in the framework region (i.e.,outside of the CDRs), and preferably has less than four, three or twoconservative amino acid substitution in the framework region.

In another preferred embodiment of the treatment method, medicaments anduses of the present invention, the PD-1 antagonist is a monoclonalantibody which specifically binds to human PD-1 and comprises (a) aheavy chain comprising SEQ ID NO: 14 and (b) a light chain comprisingSEQ ID NO:18, SEQ ID NO:19 or SEQ ID NO:20.

In yet another preferred embodiment of the treatment method, medicamentsand uses of the present invention, the PD-1 antagonist is a monoclonalantibody which specifically binds to human PD-1 and comprises (a) aheavy chain comprising SEQ ID NO: 14 and (b) a light chain comprisingSEQ ID NO:18.

Table 3 below provides a list of the amino acid sequences of exemplaryanti-PD-1 mAbs for use in the treatment method, medicaments and uses ofthe present invention, and the sequences are shown in FIGS. 1-5.

TABLE 3 Exemplary anti-human PD-1 antibodies A. Comprises light andheavy chain CDRs of hPD-1.08A in WO2008/156712 CDRL1 SEQ ID NO: 1 CDRL2SEQ ID NO: 2 CDRL3 SEQ ID NO: 3 CDRH1 SEQ ID NO: 4 CDRH2 SEQ ID NO: 5CDRH3 SEQ ID NO: 6 B. Comprises light and heavy chain CDRs of hPD-1.09Ain WO2008/156712 CDRL1 SEQ ID NO: 7 CDRL2 SEQ ID NO: 8 CDRL3 SEQ ID NO:9 CDRH1 SEQ ID NO: 10 CDRH2 SEQ ID NO: 11 CDRH3 SEQ ID NO: 12 C.Comprises the mature h109A heavy chain variable region and one of themature K09A light chain variable regions in WO2008/156712 Heavy chain VRSEQ ID NO: 13 Light chain VR SEQ ID NO: 15 or SEQ ID NO: 16 or SEQ IDNO: 17 D. Comprises the mature 409 heavy chain and one of the matureK09A light chains in WO2008/156712 Heavy chain SEQ ID NO: 14 Light chainSEQ ID NO: 18 or SEQ ID NO: 19 or SEQ ID NO: 20

“PD-L1” or “PD-L2” expression as used herein means any detectable levelof expression of the designated PD-L protein on the cell surface or ofthe designated PD-L mRNA within a cell or tissue. PD-L proteinexpression may be detected with a diagnostic PD-L antibody in an IHCassay of a tumor tissue section or by flow cytometry. Alternatively,PD-L protein expression by tumor cells may be detected by PET imaging,using a binding agent (e.g., antibody fragment, affibody and the like)that specifically binds to the desired PD-L target, e.g., PD-L1 orPD-L2. Techniques for detecting and measuring PD-L mRNA expressioninclude RT-PCR and realtime quantitative RT-PCR.

Several approaches have been described for quantifying PD-L1 proteinexpression in IHC assays of tumor tissue sections. See, e.g., Thompson,R. H., et al., PNAS 101 (49); 17174-17179 (2004); Thompson, R. H. etal., Cancer Res. 66:3381-3385 (2006); Gadiot, J., et al., Cancer117:2192-2201 (2011); Taube, J. M. et al., Sci Transl Med 4, 127ra37(2012); and Toplian, S. L. et al., New Eng. J Med. 366 (26): 2443-2454(2012).

One approach employs a simple binary end-point of positive or negativefor PD-L1 expression, with a positive result defined in terms of thepercentage of tumor cells that exhibit histologic evidence ofcell-surface membrane staining. A tumor tissue section is counted aspositive for PD-L1 expression is at least 1%, and preferably 5% of totaltumor cells.

In another approach, PD-L1 expression in the tumor tissue section isquantified in the tumor cells as well as in infiltrating immune cells,which predominantly comprise lymphocytes. The percentage of tumor cellsand infiltrating immune cells that exhibit membrane staining areseparately quantified as <5%, 5 to 9%, and then in 10% increments up to100%. For tumor cells, PD-L1 expression is counted as negative if thescore is <5% score and positive if the score is >5%. PD-L1 expression inthe immune infiltrate is reported as a semi-quantitative measurementcalled the adjusted inflammation score (AIS), which is determined bymultiplying the percent of membrane staining cells by the intensity ofthe infiltrate, which is graded as none (0), mild (score of 1, rarelymphocytes), moderate (score of 2, focal infiltration of tumor bylymphohistiocytic aggregates), or severe (score of 3, diffuseinfiltration). A tumor tissue section is counted as positive for PD-L1expression by immune infiltrates if the AIS is >5.

A tissue section from a tumor that has been stained by IHC with adiagnostic PD-L1 antibody may also be scored for PD-L1 proteinexpression by assessing PD-L1 expression in both the tumor cells andinfiltrating immune cells in the tissue section, using a novel scoringprocess described in co-pending application 61/807,581, filed 2 Apr.2013. This PD-L1 scoring process comprises examining each tumor nest inthe tissue section for staining, and assigning to the tissue section oneor both of a modified H score (MHS) and a modified proportion score(MPS). To assign the MHS, four separate percentages are estimated acrossall of the viable tumor cells and stained mononuclear inflammatory cellsin all of the examined tumor nests: (a) cells that have no staining(intensity=0), (b) weak staining (intensity=1+), (c) moderate staining(intensity=2+) and (d) strong staining (intensity=3+). A cell must haveat least partial membrane staining to be included in the weak, moderateor strong staining percentages. The estimated percentages, the sum ofwhich is 100%, are then input into the formula of 1×(percent of weakstaining cells)+2×(percent of moderate staining cells)+3×(percent ofstrong staining cells), and the result is assigned to the tissue sectionas the MHS. The MPS is assigned by estimating, across all of the viabletumor cells and stained mononuclear inflammatory cells in all of theexamined tumor nests, the percentage of cells that have at least partialmembrane staining of any intensity, and the resulting percentage isassigned to the tissue section as the MPS. In some embodiments, thetumor is designated as positive for PD-L1 expression if the MHS or theMPS is positive.

The level of PD-L mRNA expression may be compared to the mRNA expressionlevels of one or more reference genes that are frequently used inquantitative RT-PCR, such as ubiquitin C.

In some embodiments, a level of PD-L1 expression (protein and/or mRNA)by malignant cells and/or by infiltrating immune cells within a tumor isdetermined to be “overexpressed” or “elevated” based on comparison withthe level of PD-L1 expression (protein and/or mRNA) by an appropriatecontrol. For example, a control PD-L1 protein or mRNA expression levelmay be the level quantified in nonmalignant cells of the same type or ina section from a matched normal tissue. In some preferred embodiments,PD-L1 expression in a tumor sample is determined to be elevated if PD-L1protein (and/or PD-L1 mRNA) in the sample is at least 10%, 20%, or 30%greater than in the control.

“Sustained response” means a sustained therapeutic effect aftercessation of treatment with a therapeutic agent, or a combinationtherapy described herein. In some embodiments, the sustained responsehas a duration that is at least the same as the treatment duration, orat least 1.5, 2.0, 2.5 or 3 times longer than the treatment duration.

“Tissue Section” refers to a single part or piece of a tissue sample,e.g., a thin slice of tissue cut from a sample of a normal tissue or ofa tumor.

“Treat” or “treating” a cancer as used herein means to administer acombination therapy of a PD-1 antagonist and a dinaciclib compound to asubject having a cancer, or diagnosed with a cancer, to achieve at leastone positive therapeutic effect, such as for example, reduced number ofcancer cells, reduced tumor size, reduced rate of cancer cellinfiltration into peripheral organs, or reduced rate of tumor metastasisor tumor growth. Positive therapeutic effects in cancer can be measuredin a number of ways (See, W. A. Weber, J. Nucl. Med. 50:1S-10S (2009)).For example, with respect to tumor growth inhibition, according to NCIstandards, a T/C≦42% is the minimum level of anti-tumor activity. AT/C<10% is considered a high anti-tumor activity level, with T/C(%)=Median tumor volume of the treated/Median tumor volume of thecontrol×100. In some embodiments, the treatment achieved by atherapeutically effective amount is any of progression free survival(PFS), disease free survival (DFS) or overall survival (OS). PFS, alsoreferred to as “Time to Tumor Progression” indicates the length of timeduring and after treatment that the cancer does not grow, and includesthe amount of time patients have experienced a complete response or apartial response, as well as the amount of time patients haveexperienced stable disease. DFS refers to the length of time during andafter treatment that the patient remains free of disease. OS refers to aprolongation in life expectancy as compared to naive or untreatedindividuals or patients. The dosage regimen of a combination therapydescribed herein that is effective to treat a cancer patient may varyaccording to factors such as the disease state, age, and weight of thepatient, and the ability of the therapy to elicit an anti-cancerresponse in the subject. While an embodiment of the treatment method,medicaments and uses of the present invention may not be effective inachieving a positive therapeutic effect in every subject, it should doso in a statistically significant number of subjects as determined byany statistical test known in the art such as the Student's t-test, thechi²-test, the U-test according to Mann and Whitney, the Kruskal-Wallistest (H-test), Jonckheere-Terpstra-test and the Wilcoxon-test.

“Tumor” as it applies to a subject diagnosed with, or suspected ofhaving, a cancer refers to a malignant or potentially malignant neoplasmor tissue mass of any size, and includes primary tumors and secondaryneoplasms. A solid tumor is an abnormal growth or: mass of tissue thatusually does not contain cysts or liquid areas. Different types of solidtumors are named for the type of cells that form them. Examples of solidtumors are sarcomas, carcinomas, and lymphomas. Leukemias (cancers ofthe blood) generally do not form solid tumors (National CancerInstitute, Dictionary of Cancer Terms).

“Tumor burden” also referred to as “tumor load”, refers to the totalamount of tumor material distributed throughout the body. Tumor burdenrefers to the total number of cancer cells or the total size oftumor(s), throughout the body, including lymph nodes and bone narrow.Tumor burden can be determined by a variety of methods known in the art,such as, e.g. by measuring the dimensions of tumor(s) upon removal fromthe subject, e.g., using calipers, or while in the body using imagingtechniques, e.g., ultrasound, bone scan, computed tomography (CT) ormagnetic resonance imaging (MRI) scans.

The term “tumor size” refers to the total size of the tumor which can bemeasured as the length and width of a tumor. Tumor size may bedetermined by a variety of methods known in the art, such as, e.g. bymeasuring the dimensions of tumor(s) upon removal from the subject,e.g., using calipers, or while in the body using imaging techniques,e.g., bone scan, ultrasound, CT or MRI scans.

“Variable regions” or “V region” as used herein means the segment of IgGchains which is variable in sequence between different antibodies. Itextends to Kabat residue 109 in the light chain and 113 in the heavychain.

II. METHODS, USES AND MEDICAMENTS

In one aspect of the invention, the invention provides a method fortreating a cancer in an individual comprising administering to theindividual a combination therapy which comprises a PD-1 antagonist and adinaciclib compound.

The combination therapy may also comprise one or more additionaltherapeutic agents. The additional therapeutic agent may be, e.g., achemotherapeutic other than a dinaciclib compound, a biotherapeuticagent (including but not limited to antibodies to VEGF, EGFR, Her2/neu,VEGF receptors, other growth factor receptors, CD20, CD40, CD-40L,CTLA-4, OX-40, 4-1BB, and ICOS), an immunogenic agent (for example,attenuated cancerous cells, tumor antigens, antigen presenting cellssuch as dendritic cells pulsed with tumor derived antigen or nucleicacids, immune stimulating cytokines (for example, IL-2, IFNα2, GM-CSF),and cells transfected with genes encoding immune stimulating cytokinessuch as but not limited to GM-CSF).

Examples of chemotherapeutic agents include alkylating agents such asthiotepa and cyclosphosphamide; alkyl sulfonates such as busulfan,improsulfan and piposulfan; aziridines such as benzodopa, carboquone,meturedopa, and uredopa; ethylenimines and methylamelamines includingaltretamine, triethylenemelamine, trietylenephosphoramide,triethylenethiophosphoramide and trimethylolomelamine; acetogenins(especially bullatacin and bullatacinone); a camptothecin (including thesynthetic analogue topotecan); bryostatin; callystatin; CC-1065(including its adozelesin, carzelesin and bizelesin syntheticanalogues); cryptophycins (particularly cryptophycin 1 and cryptophycin8); dolastatin; duocarmycin (including the synthetic analogues, KW-2189and CBI-TMI); eleutherobin; pancratistatin; a sarcodictyin;spongistatin; nitrogen mustards such as chlorambucil, chlornaphazine,cholophosphamide, estramustine, ifosfamide, mechlorethamine,mechlorethamine oxide hydrochloride, melphalan, novembichin,phenesterine, prednimustine, trofosfamide, uracil mustard; nitrosureassuch as carmustine, chlorozotocin, fotemustine, lomustine, nimustine,ranimustine; antibiotics such as the enediyne antibiotics (e.g.calicheamicin, especially calicheamicin gamma1I and calicheamicin phiI1,see, e.g., Agnew, Chem. Intl. Ed. Engl., 33:183-186 (1994); dynemicin,including dynemicin A; bisphosphonates, such as clodronate; anesperamicin; as well as neocarzinostatin chromophore and relatedchromoprotein enediyne antibiotic chromomophores), aclacinomysins,actinomycin, authramycin, azaserine, bleomycins, cactinomycin,carabicin, caminomycin, carzinophilin, chromomycins, dactinomycin,daunorubicin, detorubicin, 6-diazo-5-oxo-L-norleucine, doxorubicin(including morpholino-doxorubicin, cyanomorpholino-doxorubicin,2-pyrrolino-doxorubicin and deoxydoxorubicin), epirubicin, esorubicin,idarubicin, marcellomycin, mitomycins such as mitomycin C, mycophenolicacid, nogalamycin, olivomycins, peplomycin, potfiromycin, puromycin,quelamycin, rodorubicin, streptonigrin, streptozocin, tubercidin,ubenimex, zinostatin, zorubicin; anti-metabolites such as methotrexateand 5-fluorouracil (5-FU); folic acid analogues such as denopterin,methotrexate, pteropterin, trimetrexate; purine analogs such asfludarabine, 6-mercaptopurine, thiamiprine, thioguanine; pyrimidineanalogs such as ancitabine, azacitidine, 6-azauridine, carmofur,cytarabine, dideoxyuridine, doxifluridine, enocitabine, floxuridine;androgens such as calusterone, dromostanolone propionate, epitiostanol,mepitiostane, testolactone; anti-adrenals such as aminoglutethimide,mitotane, trilostane; folic acid replenisher such as frolinic acid;aceglatone; aldophosphamide glycoside; aminolevulinic acid; eniluracil;amsacrine; bestrabucil; bisantrene; edatraxate; defofamine; demecolcine;diaziquone; elformithine; elliptinium acetate; an epothilone; etoglucid;gallium nitrate; hydroxyurea; lentinan; lonidamine; maytansinoids suchas maytansine and ansamitocins; mitoguazone; mitoxantrone; mopidamol;nitracrine; pentostatin; phenamet; pirarubicin; losoxantrone;podophyllinic acid; 2-ethylhydrazide; procarbazine; razoxane; rhizoxin;sizofuran; spirogermanium; tenuazonic acid; triaziquone; 2,2′,2″-trichlorotriethylamine; trichothecenes (especially T-2 toxin,verracurin A, roridin A and anguidine); urethan; vindesine; dacarbazine;mannomustine; mitobronitol; mitolactol; pipobroman; gacytosine;arabinoside (“Ara-C”); cyclophosphamide; thiotepa; taxoids, e.g.paclitaxel and doxetaxel; chlorambucil; gemcitabine; 6-thioguanine;mercaptopurine; methotrexate; platinum analogs such as cisplatin andcarboplatin; vinblastine; platinum; etoposide (VP-16); ifosfamide;mitoxantrone; vincristine; vinorelbine; novantrone; teniposide;edatrexate; daunomycin; aminopterin; xeloda; ibandronate; CPT-11;topoisomerase inhibitor RFS 2000; difluoromethylornithine (DMFO);retinoids such as retinoic acid; capecitabine; and pharmaceuticallyacceptable salts, acids or derivatives of any of the above. Alsoincluded are anti-hormonal agents that act to regulate or inhibithormone action on tumors such as anti-estrogens and selective estrogenreceptor modulators (SERMs), including, for example, tamoxifen,raloxifene, droloxifene, 4-hydroxytamoxifen, trioxifene, keoxifene,LY117018, onapristone, and toremifene (Fareston); aromatase inhibitorsthat inhibit the enzyme aromatase, which regulates estrogen productionin the adrenal glands, such as, for example, 4(5)-imidazoles,aminoglutethimide, megestrol acetate, exemestane, formestane, fadrozole,vorozole, letrozole, and anastrozole; and anti-androgens such asflutamide, nilutamide, bicalutamide, leuprolide, and goserelin; andpharmaceutically acceptable salts, acids or derivatives of any of theabove.

Each therapeutic agent in a combination therapy of the invention may beadministered either alone or in a medicament (also referred to herein asa pharmaceutical composition) which comprises the therapeutic agent andone or more pharmaceutically acceptable carriers, excipients anddiluents, according to standard pharmaceutical practice.

Each therapeutic agent in a combination therapy of the invention may beadministered simultaneously (i.e., in the same medicament), concurrently(i.e., in separate medicaments administered one right after the other inany order) or sequentially in any order. Sequential administration isparticularly useful when the therapeutic agents in the combinationtherapy are in different dosage forms (one agent is a tablet or capsuleand another agent is a sterile liquid) and/or are administered ondifferent dosing schedules, e.g., a chemotherapeutic that isadministered at least daily and a biotherapeutic that is administeredless frequently, such as once weekly, once every two weeks, or onceevery three weeks.

In some embodiments, the dinaciclib compound is administered beforeadministration of the PD-1 antagonist, while in other embodiments, thedinaciclib compound is administered after administration of the PD-1antagonist.

In some embodiments, at least one of the therapeutic agents in thecombination therapy is administered using the same dosage regimen (dose,frequency and duration of treatment) that is typically employed when theagent is used as monotherapy for treating the same cancer. In otherembodiments, the patient receives a lower total amount of at least oneof the therapeutic agents in the combination therapy than when the agentis used as monotherapy, e.g., smaller doses, less frequent doses, and/orshorter treatment duration.

Each therapeutic agent in a combination therapy of the invention can beadministered orally or parenterally, including the intravenous,intramuscular, intraperitoneal, subcutaneous, rectal, topical, andtransdermal routes of administration.

A combination therapy of the invention may be used prior to or followingsurgery to remove a tumor and may be used prior to, during or afterradiation therapy.

In some embodiments, a combination therapy of the invention isadministered to a patient who has not been previously treated with abiotherapeutic or chemotherapeutic agent, i.e., is treatment-naïve. Inother embodiments, the combination therapy is administered to a patientwho failed to achieve a sustained response after prior therapy with abiotherapeutic or chemotherapeutic agent, i.e., istreatment-experienced.

A combination therapy of the invention is typically used to treat atumor that is large enough to be found by palpation or by imagingtechniques well known in the art, such as MRI, ultrasound, or CAT scan.In some preferred embodiments, a combination therapy of the invention isused to treat an advanced stage tumor having dimensions of at leastabout 200 mm³, 300 mm³, 400 mm³, 500 mm³, 750 mm³, or up to 1000 mm³.

A combination therapy of the invention is preferably administered to ahuman patient who has a cancer that tests positive for PD-L1 expression.In some preferred embodiments, PD-L1 expression is detected using adiagnostic anti-human PD-L1 antibody, or antigen binding fragmentthereof, in an IHC assay on an FFPE or frozen tissue section of a tumorsample removed from the patient. Typically, the patient's physicianwould order a diagnostic test to determine PD-L1 expression in a tumortissue sample removed from the patient prior to initiation of treatmentwith the PD-1 antagonist and dinaciclib compound, but it is envisionedthat the physician could order the first or subsequent diagnostic testsat any time after initiation of treatment, such as for example aftercompletion of a treatment cycle.

Selecting a dosage regimen (also referred to herein as an administrationregimen) for a combination therapy of the invention depends on severalfactors, including the serum or tissue turnover rate of the entity, thelevel of symptoms, the immunogenicity of the entity, and theaccessibility of the target cells, tissue or organ in the individualbeing treated. Preferably, a dosage regimen maximizes the amount of eachtherapeutic agent delivered to the patient consistent with an acceptablelevel of side effects. Accordingly, the dose amount and dosing frequencyof each biotherapeutic and chemotherapeutic agent in the combinationdepends in part on the particular therapeutic agent, the severity of thecancer being treated, and patient characteristics. Guidance in selectingappropriate doses of antibodies, cytokines, and small molecules areavailable. See, e.g., Wawrzynczak (1996) Antibody Therapy, BiosScientific Pub. Ltd, Oxfordshire, UK; Kresina (ed.) (1991) MonoclonalAntibodies, Cytokines and Arthritis, Marcel Dekker, New York, N.Y.; Bach(ed.) (1993) Monoclonal Antibodies and Peptide Therapy in AutoimmuneDiseases, Marcel Dekker, New York, N.Y.; Baert et al. (2003) New Engl.J. Med. 348:601-608; Milgrom et al. (1999) New Engl. J. Med.341:1966-1973; Slamon et al. (2001) New Engl. J. Med. 344:783-792;Beniaminovitz et al. (2000) New Engl. J. Med. 342:613-619; Ghosh et al.(2003) New Engl. J Med. 348:24-32; Lipsky et al. (2000) New Engl. J.Med. 343:1594-1602; Physicians' Desk Reference 2003 (Physicians' DeskReference, 57th Ed); Medical Economics Company; ISBN: 1563634457; 57thedition (November 2002). Determination of the appropriate dosage regimenmay be made by the clinician, e.g., using parameters or factors known orsuspected in the art to affect treatment or predicted to affecttreatment, and will depend, for example, the patient's clinical history(e.g., previous therapy), the type and stage of the cancer to be treatedand biomarkers of response to one or more of the therapeutic agents inthe combination therapy.

Biotherapeutic agents in a combination therapy of the invention may beadministered by continuous infusion, or by doses at intervals of, e.g.,daily, every other day, three times per week, or one time each week, twoweeks, three weeks, monthly, bimonthly, etc. A total weekly dose isgenerally at least 0.05 μg/kg, 0.2 μg/kg, 0.5 μg/kg, 1 μg/kg, 10 μg/kg,100 μg/kg, 0.2 mg/kg, 1.0 mg/kg, 2.0 mg/kg, 10 mg/kg, 25 mg/kg, 50 mg/kgbody weight or more. See, e.g., Yang et al. (2003) New Engl. J. Med.349:427-434; Herold et al. (2002) New Engl. J. Med. 346:1692-1698; Liuet al. (1999) J. Neurol. Neurosurg. Psych. 67:451-456; Portielji et al.(20003) Cancer Immunol. Immunother. 52:133-144.

In some embodiments that employ an anti-human PD-1 mAb as the PD-1antagonist in the combination therapy, the dosing regimen will compriseadministering the anti-human PD-1 mAb at a dose of 1, 2, 3, 5 or 10mg/kg at intervals of about 14 days (±2 days) or about 21 days (±2 days)or about 30 days (+2 days) throughout the course of treatment.

In other embodiments that employ an anti-human PD-1 mAb as the PD-1antagonist in the combination therapy, the dosing regimen will compriseadministering the anti-human PD-1 mAb at a dose of from about 0.005mg/kg to about 10 mg/kg, with intra-patient dose escalation. In otherescalating dose embodiments, the interval between doses will beprogressively shortened, e.g., about 30 days (±2 days) between the firstand second dose, about 14 days (±2 days) between the second and thirddoses. In certain embodiments, the dosing interval will be about 14 days(±2 days), for doses subsequent to the second dose.

In certain embodiments, a subject will be administered an intravenous(IV) infusion of a medicament comprising any of the PD-1 antagonistsdescribed herein.

In one preferred embodiment of the invention, the PD-1 antagonist in thecombination therapy is nivolumab, which is administered intravenously ata dose selected from the group consisting of: 1 mg/kg Q2W, 2 mg/kg Q2W,3 mg/kg Q2W, 5 mg/kg Q2W, 10 mg Q2W, 1 mg/kg Q3W, 2 mg/kg Q3W, 3 mg/kgQ3W, 5 mg/kg Q3W, and 10 mg Q3W.

In another preferred embodiment of the invention, the PD-1 antagonist inthe combination therapy is MK-3475, which is administered in a liquidmedicament at a dose selected from the group consisting of 1 mg/kg Q2W,2 mg/kg Q2W, 3 mg/kg Q2W, 5 mg/kg Q2W, 10 mg Q2W, 1 mg/kg Q3W, 2 mg/kgQ3W, 3 mg/kg Q3W, 5 mg/kg Q3W, and 10 mg Q3W. In some particularlypreferred embodiments, MK-3475 is administered as a liquid medicamentwhich comprises 25 mg/ml MK-3475, 7% (w/v) sucrose, 0.02% (w/v)polysorbate 80 in 10 mM histidine buffer pH 5.5, and the selected doseof the medicament is administered by IV infusion over a time period of30 minutes. The optimal dose for MK-3475 in combination with dinaciclibmay be identified by dose escalation starting with 2 mg/kg and going upto 10 mg/kg with the frequency of administration matched to thatselected for dinaciclib.

In some embodiments, a liquid medicament comprising the dinaciclibcompound is infused into the individual being treated at a dose ofbetween 1 and 100 mg/m² over a time period of 1 hour to 24 hours on eachof days 1, 8 and 15 of a 28 day cycle. In some embodiments, the timeperiod for the IV infusion is 2 hours, 8 hours or 25 hours. In otherembodiments, a dinaciclib medicament is administered by a 2 hourinfusion at a dose of 50 mg/m² once every 21 days. In some embodimentsin which the cancer is CLL, the dosage regimen for the dinaciclibmedicament comprises at least two 28 day cycles: in the first cycle, thedinaciclib is administered over a 2 hour infusion at doses of 7 mg/m²,10 mg/m² and 14 mg/m² on Days 1, 8, and 15, respectively, and in thesecond and any subsequent cycles, the dinaciclib is administered on Days1, 8 and 15 at a dose of 14 mg/m² over a 2 hour infusion. In someembodiments in which the cancer is a heme malignancy or a solid tumor,the dinaciclib is delivered once every two or three weeks, and the doseachieved may include up to 50 mg/m². In some embodiments, the higheststeady state dose of up to 50 mg/m² of dinaciclib is achieved by doseescalation in 2 hour infusions separated by about 14 or 21 days.

The present invention also provides a medicament which comprises a PD-1antagonist as described above and a pharmaceutically acceptableexcipient. When the PD-1 antagonist is a biotherapeutic agent, e.g., amAb, the antagonist may be produced in CHO cells using conventional cellculture and recovery/purification technologies.

In some embodiments, a medicament comprising an anti-PD-1 antibody asthe PD-1 antagonist may be provided as a liquid formulation or preparedby reconstituting a lyophilized powder with sterile water for injectionprior to use. WO 2012/135408 describes the preparation of liquid andlyophilized medicaments comprising MK-3475 that are suitable for use inthe present invention. In some preferred embodiments, a medicamentcomprising MK-3475 is provided in a glass vial which contains about 50mg of MK-3475.

The present invention also provides a medicament which comprises adincaciclib compound and a pharmaceutically acceptable excipient. Thedinaciclib compound may be prepared as described in U.S. Pat. No.7,119,200, and may be formulated as an aqueous medicament for IVinfusion as described in WO 2009/038701. In some preferred embodiments,the dinaciclib is formulated at 5 mg/mL in a sterile, aqueous citratebuffered solution at pH 3.0 to 4.2. This medicament is stable whenstored in refrigerated conditions (2° C. to 8° C.) and protected fromlight. The sterile buffered dinaciclib solution is then diluted with0.9% Sodium Chloride Injection (250 mL) United States Pharmacopeia (USP,weight/weight) to prepare various doses for IV administration, whichshould be administered within 24 hours when stored at controlled roomtemperature (20° C. to 25° C., or 68° F. to 77° F.).

The anti-PD-1 and dinaciclib medicaments described herein may beprovided as a kit which comprises a first container and a secondcontainer and a package insert. The first container contains at leastone dose of a medicament comprising an anti-PD-1 antagonist, the secondcontainer contains at least one dose of a medicament comprising adinaciclib compound, and the package insert, or label, which comprisesinstructions for treating a patient for cancer using the medicaments.The first and second containers may be comprised of the same ordifferent shape (e.g., vials, syringes and bottles) and/or material(e.g., plastic or glass). The kit may further comprise other materialsthat may be useful in administering the medicaments, such as diluents,filters, IV bags and lines, needles and syringes. In some preferredembodiments of the kit, the anti-PD-1 antagonist is an anti-PD-1antibody and the instructions state that the medicaments are intendedfor use in treating a patient having a cancer that tests positive forPD-L1 expression by an IHC assay.

These and other aspects of the invention, including the exemplaryspecific embodiments listed below, will be apparent from the teachingscontained herein.

EXEMPLARY SPECIFIC EMBODIMENTS OF THE INVENTION

1. A method for treating a cancer in an individual comprisingadministering to the individual a combination therapy which comprises aPD-1 antagonist and a dinaciclib compound, wherein the dinaciclibcompound is the compound of Formula I

or a pharmaceutically acceptable salt of the compound of Formula I.2. A medicament comprising a PD-1 antagonist for use in combination witha dinaciclib compound for treating a cancer in an individual.3. A medicament comprising a dinaciclib compound for use in combinationwith a PD-1 antagonist for treating a cancer in an individual.4. The medicament of embodiment 3 or 4, which further comprises apharmaceutically acceptable excipient.5. Use of a PD-1 antagonist in the manufacture of medicament fortreating a cancer in an individual when administered in combination witha dinaciclib compound.6. Use of a dinaciclib compound in the manufacture of a medicament fortreating a cancer in an individual when administered in combination witha PD-1 antagonist.7. Use of a PD-1 antagonist and a dinaciclib compound in the manufactureof medicaments for treating a cancer in an individual.8. A kit which comprises a first container, a second container and apackage insert, wherein the first container comprises at least one doseof a medicament comprising an anti-PD-1 antagonist, the second containercomprises at least one dose of a medicament comprising a dinaciclibcompound, and the package insert comprises instructions for treating anindividual for cancer using the medicaments.9. The kit of embodiment 8, wherein the instructions state that themedicaments are intended for use in treating an individual having acancer that tests positive for PD-L1 expression by animmunohistochemical (IHC) assay.10. The method, medicament, use or kit of any of embodiments 1 to 9,wherein the individual is a human and the PD-1 antagonist is amonoclonal antibody, or an antigen binding fragment thereof, whichspecifically binds to human PD-L1 and blocks the binding of human PD-L1to human PD-1.11. The method, medicament, use or kit of embodiment 9, wherein the PD-1antagonist is MPDL3280A, BMS-936559, MEDI4736, MSB0010718C or amonoclonal antibody which comprises the heavy chain and light chainvariable regions of SEQ ID NO:24 and SEQ ID NO:21, respectively, ofWO2013/019906.12. The method, medicament, use or kit of any of embodiments 1 to 9,wherein the individual is a human, and the PD-1 antagonist is amonoclonal antibody, or an antigen binding fragment thereof, whichspecifically binds to human PD-1 and blocks the binding of human PD-L1to human PD-1.13. The method, medicament, use or kit of embodiment 12, wherein thePD-1 antagonist also blocks binding of human PD-L2 to human PD-1.14. The method, medicament, use or kit of embodiment 13, wherein themonoclonal antibody, or antigen binding fragment thereof, comprises: (a)light chain CDRs of SEQ ID NOs: 1, 2 and 3 and heavy chain CDRs of SEQID NOs: 4, 5 and 6; or (b) light chain CDRs of SEQ ID NOs: 7, 8 and 9and heavy chain CDRs of SEQ ID NOs: 10, 11 and 12.15. The method, medicament, use or kit of embodiment 13, wherein themonoclonal antibody, or antigen binding fragment thereof, compriseslight chain CDRs of SEQ ID NOs: 7, 8 and 9 and heavy chain CDRs of SEQID NOs: 10, 11 and 12.16. The method, medicament, use or kit of embodiment 13, wherein thePD-1 antagonist is an anti-PD-1 monoclonal antibody which comprises aheavy chain and a light chain, and wherein the heavy chain comprises SEQID NO:21 and the light chain comprises SEQ ID NO:22.17. The method, medicament, use or kit of embodiment 13, wherein thePD-1 antagonist is an anti-PD-1 monoclonal antibody which comprises aheavy chain and a light chain, and wherein the heavy chain comprises SEQID NO:23 and the light chain comprises SEQ ID NO:24.18. The method, medicament, use or kit of any of embodiments 10-17,wherein the cancer is a solid tumor.19. The method, medicament, use or kit of any of embodiments 10-17,wherein the cancer is bladder cancer, breast cancer, clear cell kidneycancer, head/neck squamous cell carcinoma, lung squamous cell carcinoma,malignant melanoma, non-small-cell lung cancer (NSCLC), ovarian cancer,pancreatic cancer, prostate cancer, renal cell cancer, small-cell lungcancer (SCLC) or triple negative breast cancer.20. The method, medicament, use or kit of any of embodiments 10-17,wherein the cancer is ipilimumab-naïve advanced melanoma and while inother preferred embodiments, the human has ipilimumab-refractoryadvanced melanoma.21. The method, medicament, use or kit of any of embodiments 10-17,wherein the cancer is a Heme malignancy.22. The method, medicament, use or kit of any of embodiments 10-17,wherein the cancer is acute lymphoblastic leukemia (ALL), acute myeloidleukemia (AML), chronic lymphocytic leukemia (CLL), chronic myeloidleukemia (CML), diffuse large B-cell lymphoma (DLBCL), follicularlymphoma, Hodgkin's lymphoma (HL), mantle cell lymphoma (MCL), multiplemyeloma (MM), myeloid cell leukemia-1 protein (Mc1-1), myelodysplasticsyndrome (MDS), non-Hodgkin's lymphoma (NHL), or small lymphocyticlymphoma (SLL).23. The method, medicament, use or kit of any of embodiments 10-17,wherein the cancer is chronic lymphocytic leukemia (CLL).24. The method, medicament, use or kit of any of embodiments 10-23, thecancer expresses human PD-L1.25. The method, medicament, use or kit of embodiment 25, the human PD-L1expression is elevated.26. The method, medicament, use or kit of embodiment 24 or 25, whereinthe human PD-L1 expression is detected using a diagnostic anti-hPD-L1mAb which comprises a light chain variable region (LCVR) and a heavychain variable region (HCVR), wherein the LCVR comprises SEQ ID NO:28and the HCVR comprises SEQ ID NO:29.27. The method, medicament, use or kit of embodiment 16, wherein thePD-1 antagonist is MK-3475 or nivolumab and the dinaciclib compound isthe compound of Formula I.28. The method, medicament, use or kit of embodiment 27, wherein theMK-3475 is formulated as a liquid medicament which comprises 25 mg/mlMK-3475, 7% (w/v) sucrose, 0.02% (w/v) polysorbate 80 in 10 mM histidinebuffer pH 5.5 and the dinaciclib compound is formulated as a liquidmedicament comprising 5 mg/mL of the compound of Formula I in a sterile,aqueous citrate buffered solution at pH 3.0 to 4.2.29. The method, medicament, use or kit of embodiment 27 or 28, whereinthe cancer is CLL.30. The method, medicament, use or kit of any of embodiments 27-29,wherein the human PD-L1 expression is detected using a diagnosticanti-hPD-L1 mAb which comprises a light chain variable region (LCVR) anda heavy chain variable region (HCVR), wherein the LCVR comprises SEQ IDNO:28 and the HCVR comprises SEQ ID NO:29.

General Methods

Standard methods in molecular biology are described Sambrook, Fritschand Maniatis (1982 & 1989 2^(nd) Edition, 2001 3^(rd) Edition) MolecularCloning, A Laboratory Manual, Cold Spring Harbor Laboratory Press, ColdSpring Harbor, N.Y.; Sambrook and Russell (2001) Molecular Cloning,3^(rd) ed., Cold Spring Harbor Laboratory Press, Cold Spring Harbor,N.Y.; Wu (1993) Recombinant DNA, Vol. 217, Academic Press, San Diego,Calif.). Standard methods also appear in Ausbel, et al. (2001) CurrentProtocols in Molecular Biology, Vols. 1-4, John Wiley and Sons, Inc. NewYork, N.Y., which describes cloning in bacterial cells and DNAmutagenesis (Vol. 1), cloning in mammalian cells and yeast (Vol. 2),glycoconjugates and protein expression (Vol. 3), and bioinformatics(Vol. 4).

Methods for protein purification including immunoprecipitation,chromatography, electrophoresis, centrifugation, and crystallization aredescribed (Coligan, et al. (2000) Current Protocols in Protein Science,Vol. 1, John Wiley and Sons, Inc., New York). Chemical analysis,chemical modification, post-translational modification, production offusion proteins, glycosylation of proteins are described (see, e.g.,Coligan, et al. (2000) Current Protocols in Protein Science, Vol. 2,John Wiley and Sons, Inc., New York; Ausubel, et al. (2001) CurrentProtocols in Molecular Biology, Vol. 3, John Wiley and Sons, Inc., NY,N.Y., pp. 16.0.5-16.22.17; Sigma-Aldrich, Co. (2001) Products for LifeScience Research, St. Louis, Mo.; pp. 45-89; Amersham Pharmacia Biotech(2001) BioDirectory, Piscataway, N.J., pp. 384-391). Production,purification, and fragmentation of polyclonal and monoclonal antibodiesare described (Coligan, et al. (2001) Current Protocols in Immunology,Vol. 1, John Wiley and Sons, Inc., New York; Harlow and Lane (1999)Using Antibodies, Cold Spring Harbor Laboratory Press, Cold SpringHarbor, N.Y.; Harlow and Lane, supra). Standard techniques forcharacterizing ligand/receptor interactions are available (see, e.g.,Coligan, et al. (2001) Current Protocols in Immunology, Vol. 4, JohnWiley, Inc., New York).

Monoclonal, polyclonal, and humanized antibodies can be prepared (see,e.g., Sheperd and Dean (eds.) (2000) Monoclonal Antibodies, Oxford Univ.Press, New York, N.Y.; Kontermann and Dubel (eds.) (2001) AntibodyEngineering, Springer-Verlag, New York; Harlow and Lane (1988)Antibodies A Laboratory Manual, Cold Spring Harbor Laboratory Press,Cold Spring Harbor, N.Y., pp. 139-243; Carpenter, et al. (2000) J.Immunol. 165:6205; He, et al. (1998) J. Immunol. 160:1029; Tang et al.(1999) J. Biol. Chem. 274:27371-27378; Baca et al. (1997) J. Biol. Chem.272:10678-10684; Chothia et al. (1989) Nature 342:877-883; Foote andWinter (1992) J. Mol. Biol. 224:487-499; U.S. Pat. No. 6,329,511).

An alternative to humanization is to use human antibody librariesdisplayed on phage or human antibody libraries in transgenic mice(Vaughan et al. (1996) Nature Biotechnol. 14:309-314; Barbas (1995)Nature Medicine 1:837-839; Mendez et al. (1997) Nature Genetics15:146-156; Hoogenboom and Chames (2000) Immunol. Today 21:371-377;Barbas et al. (2001) Phage Display: A Laboratory Manual, Cold SpringHarbor Laboratory Press, Cold Spring Harbor, N.Y.; Kay et al. (1996)Phage Display of Peptides and Proteins: A Laboratory Manual, AcademicPress, San Diego, Calif.; de Bruin et al. (1999) Nature Biotechnol.17:397-399).

Purification of antigen is not necessary for the generation ofantibodies. Animals can be immunized with cells bearing the antigen ofinterest. Splenocytes can then be isolated from the immunized animals,and the splenocytes can fused with a myeloma cell line to produce ahybridoma (see, e.g., Meyaard et al. (1997) Immunity 7:283-290; Wrightet al. (2000) Immunity 13:233-242; Preston et al., supra; Kaithamana etal. (1999) J. Immunol. 163:5157-5164).

Antibodies can be conjugated, e.g., to small drug molecules, enzymes,liposomes, polyethylene glycol (PEG). Antibodies are useful fortherapeutic, diagnostic, kit or other purposes, and include antibodiescoupled, e.g., to dyes, radioisotopes, enzymes, or metals, e.g.,colloidal gold (see, e.g., Le Doussal et al. (1991) J. Immunol.146:169-175; Gibellini et al. (1998) J. Immunol. 160:3891-3898; Hsingand Bishop (1999) J. Immunol. 162:2804-2811; Everts et al. (2002) J.Immunol. 168:883-889).

Methods for flow cytometry, including fluorescence activated cellsorting (FACS), are available (see, e.g., Owens, et al. (1994) FlowCytometry Principles for Clinical Laboratory Practice, John Wiley andSons, Hoboken, N.J.; Givan (2001) Flow Cytometry, 2^(nd) ed.;Wiley-Liss, Hoboken, N.J.; Shapiro (2003) Practical Flow Cytometry, JohnWiley and Sons, Hoboken, N.J.). Fluorescent reagents suitable formodifying nucleic acids, including nucleic acid primers and probes,polypeptides, and antibodies, for use, e.g., as diagnostic reagents, areavailable (Molecular Probesy (2003) Catalogue, Molecular Probes, Inc.,Eugene, Oreg.; Sigma-Aldrich (2003) Catalogue, St. Louis, Mo.).

Standard methods of histology of the immune system are described (see,e.g., Muller-Harmelink (ed.) (1986) Human Thymus: Histopathology andPathology, Springer Verlag, New York, N.Y.; Hiatt, et al. (2000) ColorAtlas of Histology, Lippincott, Williams, and Wilkins, Phila, Pa.;Louis, et al. (2002) Basic Histology: Text and Atlas, McGraw-Hill, NewYork, N.Y.).

Software packages and databases for determining, e.g., antigenicfragments, leader sequences, protein folding, functional domains,glycosylation sites, and sequence alignments, are available (see, e.g.,GenBank, Vector NTI® Suite (Informax, Inc, Bethesda, Md.); GCG WisconsinPackage (Accelrys, Inc., San Diego, Calif.); DeCypher® (TimeLogic Corp.,Crystal Bay, Nev.); Menne, et al. (2000) Bioinformatics 16: 741-742;Menne, et al. (2000) Bioinformatics Applications Note 16:741-742; Wren,et al. (2002) Comput. Methods Programs Biomed. 68:177-181; von Heijne(1983) Eur. J. Biochem. 133:17-21; von Heijne (1986) Nucleic Acids Res.14:4683-4690).

EXAMPLES Example 1 Anti-Tumor Response of Concurrent Administration of aPD-1 Antagonist and Dinaciclib to Tumor-Bearing Mice

This experiment compared the anti-Minor response of tumor-bearing miceto treatment with one of three regimens: monotherapy with a murineanti-mouse PD-1 monoclonal antibody (Anti-PD1), monotherapy withdinaciclib and combination therapy with these two agents administeredconcurrently.

While human tumor cells or tumor explants can be grown inimmunodeficient animals as xenografts, they cannot be used for testingimmunotherapeutics because of the lack of a functional immune system.For a meaningful evaluation of immunotherapeutics or combination ofimmunotherapeutics with other agents, it is necessary to use a syngeneicmodel in which syngeneic tumors are grown in animals with an intactimmune system. MC38 is a mouse colorectal adenocarcinoma syngeneic toC57BL/6 strain. This is a relevant model system for evaluatinganti-PD-1's mechanism of action because of the translatable molecularprofile of this tumor post-anti-PD-1 therapy.

Tumor-bearing mice for this study were initiated by implanting 1×10⁶log-phase and sub-confluent MC38 cells on the right lower dorsal flankof 8 weeks old female C57BL/6 mice with an average body weight of 20grams. When the mean tumor volume in these mice reached ˜150 cubic mm(FIG. 8B left panel marked day 0), the tumor-bearing mice wererandomized to 4 treatment groups of 12 mice per group: (1)Isotype+Vehicle control group; (2) Anti-PD1+Vehicle control; (3)dinaciclib+Isotype control and (4) Anti-PD1+dinaciclib. The Vehiclecontrol was 20% hydroxypropyl β-d-cyclodextrin made by dissolvingTrappsol (Cyclodextrin Technologies Development Inc., Alachua, Fla.) ininjection-grade water. The Isotype control was a mouse monoclonalantibody specific for adenoviral hexon and was of the isotype IgG1.Anti-PD1 was administered to treatment groups 2 and 4 at 5 mg/kg i.p.every 5 days for each of 5 cycles. Dinaciclib was administered totreatment groups 3 and 4 at 40 mg/kg every 5 days for each of 5 cycles.

As demonstrated by the results, which are shown in FIGS. 8A and 8B, themean anti-tumor response of combination therapy with the PD-1 antagonistand dinaciclib was greater (p<0.05) than the anti-tumor responseobserved with either agent as monotherapy. As shown in FIG. 8A, thecombination of these two agents provided significantly higher completeregressions (CR) such that no measureable tumor remained, compared tothe best single agent response, which was 42% CR with anti-PD-1.Comparing mean tumor volumes at the end of the study (FIG. 8B, rightpanel) using one way ANOVA, the tumor volumes of mice treated with thecombination of dinaciclib+anti-PD1 were significantly smaller than thosetreated with dinaciclib alone.

The statistical significance of the responses to the differenttreatments was determined using a Fishers Exact Pair-Wise Test, and theresults are shown in Table 4 below.

TABLE 4 Fisher's exact test: pairwise comparison of tumor volumes at theend of study Tumor Volume (day 25) Treatment Pairs Non-Zero Zero Total pvalue Isotype + Vehicle 12 0 12 0.0373 Anti-PD1 7 5 12 Total 19 5 24Dinaciclib 12 0 12 0.0373 Anti-PD1 7 5 12 Total 19 5 24 Anti-PD1 12 0 120.0046 Dinaciclib 7 5 12 Total 19 5 24 Dinaciclib 12 0 12 <0.0001Dinaciclib + Anti-PD1 0 12 12 Total 12 12 24

Table 5 provides a brief description of the sequences in the sequencelisting.

SEQ ID NO: Description 1 hPD-1.08A light chain CDR1 2 hPD-1.08A lightchain CDR2 3 hPD-1-08A light chain CDR3 4 hPD-1.08A heavy chain CDR1 5hPD-1.08A heavy chain CDR2 6 hPD-1.08A heavy chain CDR3 7 hPD-1.09Alight chain CDR1 8 hPD-1.09A light chain CDR2 9 hPD-1.09A light chainCDR3 10 hPD-1.09A heavy chain CDR1 11 hPD-1.09A heavy chain CDR2 12hPD-1.09A heavy chain CDR3 13 109A-H heavy chain variable region 14409A-H heavy chain full length 15 K09A-L-11 light chain variable region16 K09A-L-16 light chain variable region 17 K09A-L-17 light chainvariable region 18 K09A-L-11 light chain full length 19 K09A-L-16 lightchain full length 20 K09A-L-17 light chain full length 21 MK-3475 Heavychain 22 MK-3475 Light chain 23 Nivolumab Heavy chain 24 Nivolumab lightchain 25 Precursor human PD-L1

REFERENCES

-   1. Sharpe, A. H, Wherry, E. J., Ahmed R., and Freeman G. J. The    function of programmed cell death 1 and its ligands in regulating    autoimmunity and infection. Nature Immunology (2007); 8:239-245.-   2. Dong H et al. Tumor-associated B7-H1 promotes T-cell apoptosis: a    potential mechanism of immune evasion. Nat Med. 2002 August;    8(8):793-800.-   3. Yang et al. PD-1 interaction contributes to the functional    suppression of T-cell responses to human uveal melanoma cells in    vitro. Invest Ophthalmol Vis Sci. 2008 June; 49(6 (2008): 49:    2518-2525.-   4. Ghebeh et al. The B7-H1 (PD-L1) T lymphocyte-inhibitory molecule    is expressed in breast cancer patients with infiltrating ductal    carcinoma: correlation with important high-risk propgnostic factors.    Neoplasia (2006) 8: 190-198.-   5. Hamanishi J et al. Programmed cell death 1 ligand 1 and    tumor-infiltrating CD8+T lymphocytes are prognostic factors of human    ovarian cancer. Proceeding of the National Academy of Sciences    (2007): 104: 3360-3365.-   6. Thompson R H et al. Significance of B7-H1 overexpression in    kidney cancer. Clinical genitourin Cancer (2006): 5: 206-211.-   7. Nomi, T. Sho, M., Akahori, T., et al. Clinical significance and    therapeutic potential of the programmed death-1 ligand/programmed    death-1 pathway in human pancreatic cancer. Clinical Cancer Research    (2007); 13:2151-2157.-   8. Ohigashi Y et al. Clinical significance of programmed death-1    ligand-1 and programmed death-1 ligand 2 expression in human    esophageal cancer. Clin. Cancer Research (2005): 11: 2947-2953.-   9. Inman et al. PD-L1 (B7-H1) expression by urothelial carcinoma of    the bladder and BCG-induced granulomata: associations with localized    stage progression. Cancer (2007): 109: 1499-1505.-   10. Shimauchi T et al. Augmented expression of programmed death-1 in    both neoplastic and nonneoplastic CD4+ T-cells in adult T-cell    Leukemia/Lymphoma. Int. J. Cancer (2007): 121:2585-2590.-   11. Gao et al. Overexpression of PD-L1 significantly associates with    tumor aggressiveness and postoperative recurrence in human    hepatocellular carcinoma. Clinical Cancer Research (2009) 15:    971-979.-   12. Nakanishi J. Overexpression of B7-H1 (PD-L1) significantly    associates with tumor grade and postoperative prognosis in human    urothelial cancers. Cancer Immunol Immunother. (2007) 56: 1173-1182.-   13. Hino et al. Tumor cell expression of programmed cell death-1 is    a prognostic factor for malignant melanoma. Cancer (2010): 00: 1-9.-   14. Ghebeh H. Foxp3+ tregs and B7-H1+/PD-1+ T lymphocytes    co-infiltrate the tumor tissues of high-risk breast cancer patients:    implication for immunotherapy. BMC Cancer. 2008 Feb. 23; 8:57.-   15. Ahmadzadeh M et al. Tumor antigen-specific CD8 T cells    infiltrating the tumor express high levels of PD-1 and are    functionally impaired. Blood (2009) 114: 1537-1544.-   16. Thompson R H et al. PD-1 is expressed by tumor infiltrating    cells and is associated with poor outcome for patients with renal    carcinoma. Clinical Cancer Research (2007) 15: 1757-1761.-   17. Guha, M. Cyclin-dependent kinase inhibitors move into Phase III.    Nature Reviews Drug Discovery (2012) 11:892-894.    18. Parry D et al. Dinaciclib (SCH 727965), a Novel and Potent    Cyclin-Dependent Kinase Inhibitor. Molecular Cancer    Therapeutics (2010) 9: 2344-2353.

All references cited herein are incorporated by reference to the sameextent as if each individual publication, database entry (e.g. Genbanksequences or GeneID entries), patent application, or patent, wasspecifically and individually indicated to be incorporated by reference.This statement of incorporation by reference is intended by Applicants,pursuant to 37 C.F.R. §1.57(b)(1), to relate to each and everyindividual publication, database entry (e.g. Genbank sequences or GeneIDentries), patent application, or patent, each of which is clearlyidentified in compliance with 37 C.F.R. §1.57(b)(2), even if suchcitation is not immediately adjacent to a dedicated statement ofincorporation by reference. The inclusion of dedicated statements ofincorporation by reference, if any, within the specification does not inany way weaken this general statement of incorporation by reference.Citation of the references herein is not intended as an admission thatthe reference is pertinent prior art, nor does it constitute anyadmission as to the contents or date of these publications or documents.

1. A method for treating a cancer in an individual comprising administering to the individual a combination therapy which comprises an antagonist of a Programmed Death 1 protein (PD-1) and a dinaciclib compound, wherein the dinaciclib compound is the compound of Formula

or a pharmaceutically acceptable salt of the compound of Formula I.
 2. The method of claim 1, wherein the individual is a human and the PD-1 antagonist is (a) a monoclonal antibody, or an antigen binding fragment thereof, which specifically binds to human PD-1 and blocks the binding of human PD-L1 to human PD-1; or (b) a monoclonal antibody, or an antigen binding fragment thereof, which specifically binds to human PD-L1 and blocks the binding of human PD-L1 to human PD-1.
 3. The method of claim 2, wherein the PD-1 antagonist is an anti-PD-1 monoclonal antibody which comprises a heavy chain and a light chain, wherein the heavy and light chains comprise SEQ ID NO:21 and SEQ ID NO:22, respectively, or SEQ ID NO:23 and SEQ ID NO:24, respectively.
 4. The method of any of claims 1 to 3, wherein the cancer is a solid tumor.
 5. The method of any of claims 1 to 3, wherein the cancer is a Heme malignancy.
 6. The method of any of claims 3 to 5, wherein the PD-1 antagonist is MK-3475 and the dinaciclib compound is the compound of Formula
 1. 7. A medicament comprising an antagonist of a Programmed Death 1 protein (PD-1) for use in combination with a dinaciclib compound for treating a cancer in an individual, wherein the dinaciclib compound is the compound of Formula I

or a pharmaceutically acceptable salt of the compound of Formula I.
 8. A medicament comprising a dinaciclib compound for use in combination with an antagonist of a Programmed Death 1 protein (PD-1) for treating a cancer in an individual, wherein the dinaciclib compound is the compound of Formula I


9. The medicament of claim 7 or 8, wherein the individual is a human and the PD-1 antagonist is (a) a monoclonal antibody, or an antigen binding fragment thereof, which specifically binds to human PD-1 and blocks the binding of human PD-L1 to human PD-1; or (b) a monoclonal antibody, or an antigen binding fragment thereof, which specifically binds to human PD-L1 and blocks the binding of human PD-L1 to human PD-1.
 10. The medicament of any of claims 7 to 9, wherein the PD-1 antagonist is an anti-PD-1 monoclonal antibody which comprises a heavy chain and a light chain, wherein the heavy and light chains comprise SEQ ID NO:21 and SEQ ID NO:22, respectively, or SEQ ID NO:23 and SEQ ID NO:24, respectively.
 11. The medicament of any of claims 7 to 10, wherein the cancer is a solid tumor.
 12. The medicament of any of claims 7 to 11, wherein the cancer is a Heme malignancy.
 13. The medicament of any of claims 10 to 12, wherein the PD-1 antagonist is MK-3475 or nivolumab.
 14. The medicament of claim 13, wherein the MK-3475 is formulated as a liquid medicament which comprises 25 mg/ml MK-3475, 7% (w/v) sucrose, 0.02% (w/v) polysorbate 80 in 10 mM histidine buffer pH 5.5 and the dinaciclib compound is formulated as a liquid medicament comprising 5 mg/mL of the compound of Formula I in a sterile, aqueous citrate buffered solution at pH 3.0 to 4.2.
 15. The medicament of claim 8, wherein the dinaciclib compound is formulated as a liquid medicament comprising 5 mg/mL of the compound of Formula I in a sterile, aqueous citrate buffered solution at pH 3.0 to 4.2.
 16. A kit which comprises a first container, a second container and a package insert, wherein the first container comprises at least one dose of a medicament comprising an antagonist of a Programmed Death 1 protein (PD-1), the second container comprises at least one dose of a medicament comprising a dinaciclib compound, and the package insert comprises instructions for treating an individual for cancer using the medicaments, wherein the dinaciclib compound is the compound of Formula I


17. The kit of claim 16, wherein the instructions state that the medicaments are intended for use in treating an individual having a cancer that tests positive for PD-L1 expression by an immunohistochemical (IHC) assay.
 18. The kit of claim 16 or 17, wherein the individual is a human and the PD-1 antagonist is (a) a monoclonal antibody, or an antigen binding fragment thereof, which specifically binds to human PD-1 and blocks the binding of human PD-L1 to human PD-1; or (b) a monoclonal antibody, or an antigen binding fragment thereof, which specifically binds to human PD-L1 and blocks the binding of human PD-L1 to human PD-1.
 19. The kit of any of claims 16-18, wherein the PD-1 antagonist is MK-3475.
 20. The method, use or kit of any of claims 1 to 19, wherein the cancer is bladder cancer, breast cancer, clear cell kidney cancer, head/neck squamous cell carcinoma, lung squamous cell carcinoma, malignant melanoma, non-small-cell lung cancer (NSCLC), ovarian cancer, pancreatic cancer, prostate cancer, renal cell cancer, small-cell lung cancer (SCLC), triple negative breast cancer, acute lymphoblastic leukemia (ALL), acute myeloid leukemia (AML), chronic lymphocytic leukemia (CLL), chronic myeloid leukemia (CML), diffuse large B-cell lymphoma (DLBCL), follicular lymphoma, Hodgkin's lymphoma (HL), mantle cell lymphoma (MCL), multiple myeloma (MM), myeloid cell leukemia-1 protein (Mc1-1), myelodysplastic syndrome (MDS), non-Hodgkin's lymphoma (NHL), or small lymphocytic lymphoma (SLL). 